Abstract Introduction: The mammalian target of rapamycin (mTOR), a downstream kinase of the PI3K/Akt signaling pathway, is a critical regulator of basic cellular functions, especially in tumor progression, and therefore has emerged as a key target for cancer chemotherapy. Rapamycin and its derivatives (e.g. Everolimus, Temsirolimus and Deferolimus) are mTOR inhibitors originally approved as immunosuppressants to prevent renal and cardiac allograft rejection. These mTOR inhibitors have recently been used in a large number of cancer clinical trials, including renal cell carcinoma, breast cancer, endometrial carcinoma, glioblastoma and mantle cell lymphoma. However, severe side effects such as toxicity and immune suppression limit the escalation of mTOR inhibitors' dosage, resulting in a low responsive rate. To maximize the efficacy and minimize side effects of mTOR inhibitors, there is a critical need to identify the genetic biomarkers for response to mTOR inhibitors and the mechanisms by which they manifest their therapeutic benefits in cancer. Purpose: This study aims to identify and validate novel genetic biomarkers that might influence the therapeutic responses of mTOR inhibitors. Methods: We used 273 human lymphoblastoid cell lines (LCLs), for which we have obtained genome-wide SNP data, Affymetric U133 plus 2 mRNA expression array data, and Illumina microRNA array data, to perform cytotoxicity assays with two mTOR inhibitors, Rapamycin and Everolimus. We then performed genome-wide association (GWA) analyses using SNPs, mRNA gene expression and microRNA data with cytotoxicity data (AUC values) collected on both mTOR inhibitors. Integrated analysis was then performed to identify the SNPs, and microRNAs that might be associated with genes, whose expressions in turn were significantly associated with the cytotoxicity. We then selected candidate genes to functionally validate their effects on sensitivity of mTOR inhibitors using an siRNA screening approach with 3 cell lines, a renal carcinoma (Caki2), a glioblastoma (U87) and a fibroblast (IMR90) cell line, followed by MTS cell proliferation assay and colony formation assay. Results: mRNA expression levels of 48 and 55 genes were associated with Rapamycin and Everolimus cytotoxicity (p<10−4) respectively, with 16 common genes between two drugs. One hundred and twenty seven and 100 SNPs were associated with Rapamycin and Everolimus cytotoxicity (p<10−4), respectively. We selected 23 candidate genes based on the GWA analyses (mRNA expression vs. cytotoxicity, SNPs vs. cytotoxicity and integrated analysis), and verified 13 genes that significantly altered cells' sensitivity to either one or both mTOR inhibitors in at least one cell line based on siRNA screening study. In addition, one microRNA, miR10a, was shown to be most significantly associated with cytotoxicity for both Rapamycin and Everolimus (p=3.3×10−4; p =1.4×10−4). Interestingly, this particular microRNA was also found to be strongly associated with gene expression levels of 9 out 23 selected candidate genes with p<10−4. Conclusions: This study identified a series of novel candidate genes and microRNA (miR10a) that might contribute to the response of mTOR inhibitors. These findings will enhance our understanding of the regulation of mTOR pathway and the mechanisms underlying the variation in response to mTOR inhibitors. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2011 Nov 12-16; San Francisco, CA. Philadelphia (PA): AACR; Mol Cancer Ther 2011;10(11 Suppl):Abstract nr PR-11.
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Jul 22, 2022
Huei-Chung Huang + 3
Open Access
